[Objectives] The objective of this study is to optimize the fermentation conditions of Bacillus subtilis NTGB-178 for improving the liquid fermentation of biomass and the spore yield, then provide a cost-effective fermentation technique.[Methods] Response surface methodology (RSM)was used to optimize the fermentation conditions. The main factors which affected the liquid fermentation were screened by Plackett-Burman design, based on the single factor experiment. The steepest ascent path was used to identify the central point of response surface and approach the optimal region. The Box-Behnken design was adopted to identify the optimal fermentation conditions by establishing a multiple quadratic regression equation, analyzing and evaluating the factors which affect the spore yield of B. subtilis NTGB-178.[Results] Bran, NaCl and initial pH were identified as the main factors. The optimum conditions, which were predicted by the response surface equation, were bran 10.35 g·L-1, NaCl 4.41 g·L-1, and initial pH6.07. The other factors were as follows:corn flour 12.5 g·L-1, bean pulp 25.0 g·L-1, CaCO31.0 g·L-1, MgSO4·7H2O 2.0 g·L-1, inoculation amount 4%, loaded liquid volume 65 mL/250 mL, rotation speed 220 r·min-1and cultivating at 36℃ for 36 h. After verification, the biomass reached 6.55×109 CFU·mL-1, and the spore yield reached 6.16×109 CFU·mL-1, which was improved 8.48 times compared to the result before optimization. In the 100 L fermenter enlarge cultivations, the spore yield was 7.33×109 CFU·mL-1. The stability of the optimal fermentation conditions wase indicated.[Conclusions] Single factor experiment and RSM were simultaneously used in the optimization of fermentation of B. subtilis NTGB-178 for the biomass and spore production, which was proved to be effective. This study laid the foundation for industrialized production of B. subtilis NTGB-178.